NANOGENESEQ. technologies address these two needs with a highly innovative and efficient system which optimize DNA sequencing at both the efficiency and cost level.There are various methods to sequence DNA. The most relevant one are based on electrophoresis, capillary action, capillary gel electrophoresis, ion separation, phase transition etc. The most widely used are based on capillary electrophoresis separation of DNA strands followed by detection, again by various methods.Capillary electrophoresis separation involves flowing different molecular weight or different size DNA segments through a porous or gel material which allows smooth and easy movement in the presence of an electric field. The DNA segments are obtained by the use of several restriction enzymes which, acting each as a different chemical scissor, are able to cut the DNA at a specific site along its sequence. The flow patterns, which include the different DNA segments, are then plotted on a particular material in time and scale and the segments are detected using usually lasers (or other detection methods) in combination with certain base-specific fluorescent dyes. The founders are now seeking qualified Venture Capital to back their licensing efforts and to execute on the plan.

The core technology

NANOGENESEQ has designed an innovative sequencing system design consisting of two key components:

a) The Hardware component b) The Software componentHardware

The hardware component of NANOGENESEQ sequencing system is the most critical component since it speeds of the process of DNA sequencing altogether. It has been designed as a silicon chip in which DNA segment separation and detection is much more efficient than currently-used electrophoretic methods.DNA Separation solutions

Separating DNA fragments by length is the single most important step of the sequencing process. The traditional method is to place a “restriction enzyme digested” sample at one end of a column of an organic gel and apply an electric field to the column. The electrical field causes the flow of the DNA fragments through the gel and the rate of flow of each fragment is directly related to its molecular weight. As they slowly make their way through the tiny pores of the material, fragments of different lengths moves at different speeds and eventually collect in a series of bands as a ladder like structure that can be photographed using fluorescent or radioactive tags. One of the major problems with this conventional scheme is the overlapping of DNA fragments caused by the intense density of the media (gel or membrane) acting as a “viscous” substrate through which the fragments move.DNA fragment overlapping is the first problem that NANOGENESEQ system eliminates. NANOGENESEQ has designed NANOCHIP as a microelectronic device , a chip-like system , which allows DNA fragments to travel freely without intense density filling . It has been designed through the use of fractal geometrical methods .They are genetic algorithms which can be modelled) to develop a porous geometry architecture which allows DNA samples to move in non-viscous media , such as water and other low/zero resistance media, using an electric field. In conventional schemes the second rate-limiting step is that only a few DNA fragments can be flown through the columns to avoid overlapping. Using the porous geometry architecture has NANOGENESEQ designed a system which allows to move many fragments of different lengths at the same time, thus avoiding the overlapping problem discussed above, and at a higher speed. Recent work has shown that when DNA samples are placed in a water solution and are flown through channels, consisting of deep and shallow alternating areas, by applying an electric field to draw the DNA fragments towards the other end of the channel, what happens is that in the deep section the chain-like DNA fragments contract into a roughly spherical shape, too large to fit through the shallow sections. Hence, at the border between a deep and a shallow section, the fragment is delayed until it can stretch out thin enough to fit through the shallow section of the channel. When this happens the large fragment zips through the shallow channel section faster than short segments, the opposite of what happens in organic gels. The reason is that the longer strand in the form of a large sphere presses a larger area against the barrier, meaning that there are more parts of itself that extend into the shallow area to form a sort of beachhead which pulls the rest of the fragment through. Based on this observation we have designed a proprietary architecture silicon device with channels made up of 2400 sections, each about 1.5 micrometer (1 millionth of a meter) deep, separated by shallow barrier section of 90 – 120 nanometer (billionths of a meter ) deep, with a specific size and geometry. When DNA samples migrate through this device, fragments of different lengths travel a different speeds and arrive at the end in separate bands.In this device, due to this highly innovative architecture of the channels, the flow of the DNA takes a well definined and predictable pattern: the larger fragments move faster and are followed by successive groups of progressively thinner fragments . Fragments having a similar molecular weight arrive more or less together. At the end of the channels, the DNA strands are tagged with fluorescent materials and the peaks of fluorescent and the end of the channel measured over time forms the readout. A key advantage of NANOGENESEQ NANOCHIP device is that all the long strands can be collapsed to a spherical form, thus promoting separation of very long strands of DNA fragments. Conventional methods, employing organic gels are unable to do so, since DNA fragments in organic gels do not collapse into a spherical form.Another significant advantage (time & cost-wise) that NANOGENESEQ device offers is that it makes it easy to extract separate DNA for further analysis. As a band of strands of a particular length arrives at the end of the channel it is easily syphoned off, being a water solution. Conversely, extraction of DNA samples from an organic gel involves a complex, time-consuming and costly chemical separation process.Detection solutionsThe second important solution that NANOGENESEQ system offers with respect to conventional systems is in the second most important step of DNA sequencing, i.e. detection.

Conventional systems use lasers. The dyes used are low molecular weight fluorescent dyes. Lasers distort low molecular weight dyes and severely restrict their detection, hence, low molecular weight dyes are relatively inefficient as a detection system for DNA fragments. In fact, the argon ion laser (which is the excitation source in many flow cytometers, confocal laser scanning microscopes, laser scanners, sequencers etc. ) the wavelengths used to excite green, yellow, orange, red fluorescent dyes are limited primarily due to the 488 and 514 nm spectral lines. This severely restricts simultaneous multicolor detection. So only a very few low molecular weight dyes with fluorescent properties can be used. This is a significant unsolved problem that conventional systems are faced with.To offer a solution to this problem, NANOGENESEQ has designed a laser detection configuration which greatly reduces this problem. In conventional systems the most that can be achieved in terms of base pair readout is 600-700 base pairs for any DNA fragment strechs. This limitation is due to the fact that conventional schemes use a linear 2-D (ladder like) approach where the overlapping problem is the rate limiting factor. NANOGENESEQ system, conversely, identifies the fluorescent labeled fragments and reads them by using a spiral scheme which provides almost 3 times more data available to read. The detection and read-out efficiency is thus increased three-fold to 1800 - 2100 base pairs for any DNA fragment strechs. In addition this system avoids the severe restriction imposed on multi colour detection mentioned earlier .This is a very new innovative methods as no other relevant methods are known. In conclusion, the hearth of NANOGENESEQ system is composed of two innovative hardware components, the separation and the detection devices. The Making this two units function as a single device just like the recording device makes the pro-type DNA Braid Generator. The software is then developed according to the scheme, there are some software already available, with a bit of alteration it can be made suitable for our purposes.

Software

The proprietary software that has NANOGENESEQ Developed using fractal imaging techniques, relates to the novel porous geometrical architecture that is ta the hearth of the NANOCHIP device. The additional software needed for detection read-out purposes is commecially available and can be readily adapted to the system’s needs.